Semenogelin I (Sg I) and the fragments of peptides hydrolyzed from Sg I by prostate-specific antigen have multiple biological activities. There exists a controversy over the inhibitory effect of the key fragment on sperm motility. This article focuses on the sperm-inhibiting and antibacterial activities of the fragments of Sg I-derived peptides and illustrates the supposition concerning the most controversial aspect. A deeper insight into the action mechanisms of Sg I-derived peptides may help improve the methods of sperm screening and provide a new perspective in the management of asthenozoospermia and urinary tract infection.
Anti-Bacterial Agents ; Humans ; Male ; Semen ; drug effects ; Seminal Vesicle Secretory Proteins ; genetics ; physiology ; Spermatozoa ; drug effects

Semen liquefaction and sperm capacitation are the key processes for sperm to acquire forward movement ability. In these processes, semenogelin plays a vital role by directly participating in the formation of semen coagulation, collaborating with other protease and metal ions from the male reproductive tract, and then reacting with the surface of sperm cells, finally involved in the regulation of these processes and ensuring sperm's acquisition of forward movement ability.
Humans ; Male ; Semen ; chemistry ; Seminal Vesicle Secretory Proteins ; physiology ; Sperm Motility

Sperm acquires capacity of motility and fertility during the process of semen coagulation and liquefaction. The main coagulative protein is Semenogelin I (Sg I), specifically produced by seminal vesicles, and then decomposed by prostate specific antigens (PSA) in sperm liquefaction into a series of small fragments. These fragments, with a variety of physiological functions, are very important for the regulation of sperm capacity acquisition and progressive movement.
Humans ; Male ; Seminal Vesicle Secretory Proteins ; physiology ; Seminal Vesicles ; metabolism ; physiology

AIMTo study the molecular mechanism of epididymal protease inhibitor (Eppin) modulating the process of prostate specific antigen (PSA) digesting semenogelin (Sg).

METHODSHuman Sg cDNA (nucleotides 82-849) and Eppin cDNA (nucleotides 70-723) were generated by polymerase chain reaction (PCR) and cloned into pET-100D/TOPO. Recombinant Eppin and Sg (rEppin and rSg) were produced by BL21 (DE3). The association of Eppin with Sg was studied by far-western immunoblot and radioautography. In vitro the digestion of rSg by PSA in the presence or absence of rEppin was studied. The effect of anti-Q20E (N-terminal) and C-terminal of Eppin on Eppin-Sg binding was monitored.

RESULTSEppin binds Sg on the surface of human spermatozoa with the C-terminal of Eppin (amino acids 75-133). rSg was digested with PSA and many low molecular weight fragments were produced. When rEppin is bound to rSg, then digested by PSA, incomplete digestion and a 15-kDa fragment results. Antibody binding to the N-terminal of rEppin did not affect rSg digestion. Addition of antibodies to the C-terminal of rEppin inhibited the modulating effect of rEppin.

CONCLUSIONEppin protects a 15-kDa fragment of rSg from hydrolysis by PSA.

Animals ; Antibodies ; pharmacology ; Autoradiography ; Humans ; Hydrolysis ; Male ; Prostate-Specific Antigen ; metabolism ; Proteinase Inhibitory Proteins, Secretory ; genetics ; immunology ; metabolism ; Rabbits ; Recombinant Proteins ; genetics ; metabolism ; Semen ; cytology ; metabolism ; Seminal Vesicle Secretory Proteins ; metabolism ; Spermatozoa ; metabolism

We have previously shown that seminal vesicle protein IV (SV-IV) and its 1-70 N-terminal fragment have anti-inflammatory activity and modulate anti-thrombin III (AT) activity. Moreover, mass spectrometry analysis of purified SV-IV has shown that the protein was found to be highly heterogeneous and 14% of the total SV-IV molecules are truncated forms, of particular interest the 1-16, 1-17, and 1-18 peptides. In this work we report experimental data which demonstrate that the 1-16 peptide (P1-16) possesses a marked effect on the AT activity by preventing the formation of the thrombin-AT complex. We found that the formation of thrombin-AT complex is markedly decreased in the presence of P1-16 used at equimolar concentration with thrombin as evaluated with SDS-PAGE. We also monitored the conformational changes of thrombin in the presence of different P1-16 concentrations, and calculated the K(d) of thrombin/P1-16 system by circular dichroism technique. The probable interaction sites of P1-16 with thrombin have been also evaluated by molecular graphics and computational analyses. These results have potential implications in the treatment of sterility and thrombotic diseases.
Amino Acid Sequence ; Animals ; Antithrombin III/metabolism ; Blood Coagulation/drug effects ; Circular Dichroism ; Humans ; Models, Molecular ; Molecular Sequence Data ; Peptide Fragments/*chemistry/pharmacology ; Protein Binding/drug effects ; Protein Structure, Secondary ; Protein Structure, Tertiary ; Rats ; Seminal Vesicle Secretory Proteins/*chemistry/genetics/metabolism ; Thrombin/*chemistry/genetics/metabolism

OBJECTIVETo study the inhibition activity of Semenogelin (Sg) and its different peptides to human spermatozoa.

METHODSHuman Sg DNA and its N-terminal Sg and C-terminal Sg DNA were cloned into PET-100 vector. Positive colonies were screened and transformed into E. Coli BL21 (DE3). Recombinant Sg and its peptides were induced and expressed in high competent E. coli BL21 (DE3) , and purified by 50% Ni-NTA column. Inhibition activity assay was done by adding 4 different concentrations of semenogelin and its two peptides, 0, 1, 5 and 10 ng/microl, to human spermatozoa.

RESULTSThe peptide of Semenogelin that inhibits the activity of human spermatozoa was located in its N-terminal fragment. C-terminal Sg did not inhibit the activity of spermatozoa.

CONCLUSIONN-terminal Sg is the inhibition peptide of the whole molecular Sg. During semen liquefaction, this peptide should be cut off from the surface of human spermatozoa before they move forward.

Cloning, Molecular ; Dose-Response Relationship, Drug ; Escherichia coli ; genetics ; Humans ; Recombinant Proteins ; biosynthesis ; pharmacology ; Seminal Vesicle Secretory Proteins ; biosynthesis ; genetics ; pharmacology ; Sperm Motility ; drug effects

The epididymal protease inhibitor (Eppin) abounds in human semen and on the surface of human spermatozoa, specifically produced by the testis and epididymis. Recombinant Eppin has effected infertility in the immunized monkey and promises to be an effective vaccine for human immunocontraception. This article reviews the advances in the studies of Eppin gene and protein construction and its molecular mechanism of causing immunologic infertility and regulating PSA hydrolysis of Semenogelin.
Animals ; Humans ; Infertility, Male ; immunology ; Male ; Mice ; Primates ; Proteinase Inhibitory Proteins, Secretory ; chemistry ; genetics ; immunology ; physiology ; Seminal Vesicle Secretory Proteins ; physiology ; Vaccines, Contraceptive

OBJECTIVETo produce recombinant human prostate-specific antigen (PSA) by molecular cloning technology and to identify its activity.

METHODSThe human PSA cDNA and PET-12a vector were digested by NdeI and BamH1 before ligated by T4 ligase. The correct sequence was verified and transformed into high competent E. coli BL21 (DE3). Recombinant PSA was expressed and purified by hydrophobic interaction phenyl Sepharose column and activated by trypsin digestion. Enzymatic activation assay was done by hydrolysis of the substrate S-2586 and semenogelin.

RESULTSNon-active recombinant PSA was digested by trypsin and demonstrated enzyme activity. The activated PSA hydrolyzed S-2586 and its physiological substrate semenogelin (Sg).

CONCLUSIONRecombinant pro-PSA can be an active serine protease by trypsin digestion and demonstrate native PSA enzymatic activity.

Blotting, Western ; Cloning, Molecular ; DNA, Complementary ; genetics ; Escherichia coli ; genetics ; Gene Expression ; Humans ; Hydrolysis ; Male ; Oligopeptides ; metabolism ; Prostate-Specific Antigen ; genetics ; isolation & purification ; metabolism ; Recombinant Proteins ; isolation & purification ; metabolism ; Seminal Vesicle Secretory Proteins ; metabolism ; Trypsin ; metabolism

Sperm membrane proteins play a key role in spermatogenesis, sperm maturation and sperm-oocyte interaction. A deeper research would shed new light on the molecular mechanisms of spermatogenesis, sperm maturation and fertilization. In recent years, with the extensive application of a variety of current molecular biological methods and bioinformatics to reproductive medicine, some sperm membrane proteins found previously have been cloned and sequenced. Furthermore, new sperm membrane proteins, being found continuously, will make a solid foundation for the development of contraceptive vaccine as well as for the investigation into the mechanism of fertilization at levels of gene and protein. This article reviews the current progress in the researches on sperm membrane proteins.
Adaptor Proteins, Signal Transducing ; physiology ; Animals ; Humans ; Male ; Membrane Proteins ; physiology ; Seminal Vesicle Secretory Proteins ; physiology ; Spermatogenesis ; physiology ; Spermatozoa ; cytology ; growth & development

OBJECTIVETo investigate the binding of secretory proteins in the ventral prostate to the surface of sperm.

METHODSWe used different techniques to demonstrate the possibility of ventral prostate secretory proteins binding to sperm in golden hamsters. Polyclonal antibodies against crude secretion of the ventral prostate cultured in rabbits were used to detect the antigens in hamster epididymal, uterine and oviductal spermatozoa by indirect immunofluorescence technique. The uterine and oviductal spermatozoa were collected after mating with the males with or without ventral prostate glands. The ventral prostate secretory proteins were isolated and transblotted to the membrane, which was incubated with the biotinylated epididymal sperm membrane proteins, and then the biotinylated binding proteins were stained.

RESULTSAn immunoreaction restricted to the middle piece was observed in the sperm incubated with the ventral prostate secretion and ejaculated sperm recovered from the uteri and oviducts. The rate of the epididymal sperm bound with the ventral prostate secretory proteins was (80 +/- 5) %, and the rats of the sperm binding to the ventral prostate secretory proteins were (30.0 +/- 4.6) % from the uterus and (16.0 +/- 3.6) % from the oviduct after mating with the males with ventral prostate glands, significantly higher than after mating with those without prostate glands (P < 0.01). Five bands were identified by Western blot analysis in vitro of the ventral prostate secretory proteins incubated with biotinylated epididymal sperm membrane proteins.

CONCLUSIONThe present data indicate that ventral prostate secretory proteins bind to the middle piece of sperm in golden hamsters.

Animals ; Blotting, Western ; Cricetinae ; Epididymis ; metabolism ; Fallopian Tubes ; metabolism ; Female ; Fluorescent Antibody Technique, Indirect ; Male ; Mesocricetus ; Prostate ; metabolism ; Protein Binding ; Seminal Vesicle Secretory Proteins ; metabolism ; Spermatozoa ; metabolism ; Uterus ; metabolism